The interaction between catecholamines and insulin in regulating glucose transport in isolated rat adipose cells has been evaluated. The initial results suggest that catecholamines counterregulate insulin-stimulated glucose transport in rat adipose cells through a cAMP-mediated mechanism, but only in part by modulating the translocation of glucose transporters. The mechanism of the counterregulatory effects of lipolytic and antilipolytic hormones on insulin-stimulated 3-0-methylglucose transport in rat adipose cells has been further investigated under the stringent incubation conditions described by Londos et al. (Adv. Cyc. Nuc. Res. 17) that maintain steady state A-kinase activity ratios (plus/minus cAMP), a measure of cAMP concentration. Under these conditions, lipolytic and antilipolytic hormones counterregulate insulin stimulated glucose transport activity through a cAMP-independent modulation of glucose transporter intrinsic activity and a marked change in sensitivity to insulin apparently occurring at the level of the signaling mechanism.